Project description:Small RNAs (21-24 nt) are pivotal regulators of gene expression that guide both transcriptional and post-transcriptional silencing mechanisms in diverse eukaryotes, including most if not all plants. MicroRNAs (miRNAs) and short interfering RNAs (siRNAs) are the two major types, both of which have a demonstrated and important role in plant development, stress responses and pathogen resistance. In this work, we used a deep sequencing approach (Sequencing-By-Synthesis, or SBS) to develop sequence resources of small RNAs from Panicum virgatum tissues (including leaves, drought-treated leaves and flowers). The high depth of the resulting datasets enabled us to examine in detail critical small RNA features as size distribution, tissue-specific regulation and sequence conservation between different organs in this species. We also developed database resources and a dedicated website (http://smallrna.udel.edu/) with computational tools for allowing other users to identify new miRNAs or siRNAs involved in specific regulatory pathways, verify the degree of conservation of these sequences in other plant species and map small RNAs on genes or larger regions of the maize genome under study. Small RNA libraries were derived from leaves, drought-treated leaves and flowers of Panicum virgatum. Total RNA was isolated using the Plant RNA Purification Reagent (Invitrogen), and submitted to Illumina (Hayward, CA, http://www.illumina.com) for small RNA library construction using approaches described in (Lu et al., 2007) with minor modifications. The small RNA libraries were sequenced with the Sequencing-By-Synthesis (SBS) technology by Illumina. PERL scripts were designed to remove the adapter sequences and determine the abundance of each distinct small RNA. We thank Pamela Green for providing the plant material as well as Kan Nobuta and Gayathri Mahalingam for assistance with the computational methods.
Project description:Small RNAs (21-24 nt) are pivotal regulators of gene expression that guide both transcriptional and post-transcriptional silencing mechanisms in diverse eukaryotes, including most if not all plants. MicroRNAs (miRNAs) and short interfering RNAs (siRNAs) are the two major types, both of which have a demonstrated and important role in plant development, stress responses and pathogen resistance. In this work, we used a deep sequencing approach (Sequencing-By-Synthesis, or SBS) to develop sequence resources of small RNAs from Panicum virgatum tissues (including leaves, drought-treated leaves and flowers). The high depth of the resulting datasets enabled us to examine in detail critical small RNA features as size distribution, tissue-specific regulation and sequence conservation between different organs in this species. We also developed database resources and a dedicated website (http://smallrna.udel.edu/) with computational tools for allowing other users to identify new miRNAs or siRNAs involved in specific regulatory pathways, verify the degree of conservation of these sequences in other plant species and map small RNAs on genes or larger regions of the maize genome under study.
Project description:Cereal aphids can successfully colonize and damage switchgrass (Panicum virgatum) plants. Among the aphids tested, greenbugs (Schizaphis graminum, GB) caused significant plant damage likely through a combination of aphid-salivary proteins that are injected into plants during feeding and a strong host response elicited by herbivory. In this study, changes in protein phosphorylation present in GB-infested and uninfested control plants was determined. These data were compared against transcriptome changes recently published for this system.
Project description:Cereal aphids can successfully colonize and damage switchgrass (Panicum virgatum) plants. Among the aphids tested, greenbugs (Schizaphis graminum, GB) caused significant plant damage likely through a combination of aphid-salivary proteins that are injected into plants during feeding and a strong host response elicited by herbivory. In this study, shotgun label-free proteomics has been used to document changes to the switchgrass proteome as a result of GB infestation. These proteomic data were compared against transcriptome changes recently published for this system.
Project description:Sustainable production of switchgrass (Panicum virgatum) as a bioenergy crop hinges in part on efficient use of soil macronutrients, especially nitrogen (N). This study investigated the physiological, metabolic and transcriptomic responses of switchgrass to N limitation. Moderate N limitation marked a tipping point for large changes in plant growth, root-to-shoot ratio, root system architecture and total nitrogen content. Integration of transcriptomic and metabolic data revealed that N limitation reduced switchgrass photosynthetic capacity and carbon(C)-fixation activities. Switchgrass balanced C-fixation with N-assimilation, transport and recycling of N compounds by rerouting C-flux from glycolysis, the oxidative branch of the pentose phosphate pathway (OPPP) and from the tricarboxylic acid (TCA) cycle in an organ specific manner. The energy and reduction power so generated, and C-skeletons appear to be directed towards N uptake, biosynthesis of energy storage compounds with high C/N ratio such as sucrose, non-N-containing lipids, and various branches of secondary metabolism.
Project description:In light of the changes in precipitation and soil water availability expected with climate change, understanding the mechanisms underlying plant responses to water deficit is essential. Toward that end we have conducted an integrative analysis of responses to drought stress in the perennial C4 grass and biofuel crop, Panicum virgatum (switchgrass). Responses to soil drying and re-watering were measured at transcriptional, physiological, and metabolomic levels. To assess the interaction of soil moisture with diel light:dark cycles, we profiled gene expression in drought and control treatments under pre-dawn and mid-day conditions. Soil drying resulted in reduced leaf water potential, gas exchange, and chlorophyll fluorescence along with differential expression of a large fraction of the transcriptome (37%). Many transcripts responded differently depending on time of day (e.g. up-regulation pre-dawn and down-regulation mid-day). Genes associated with C4 photosynthesis were down-regulated during drought, while C4 metabolic intermediates accumulated. Rapid changes in gene expression were observed during recovery from drought, along with increased water use efficiency and chlorophyll fluorescence. Our findings demonstrate that drought responsive gene expression depends strongly on time of day and that gene expression is extensively modified during the first few hours of drought recovery. Analysis of covariation in gene expression, metabolite abundance, and physiology among plants revealed non-linear relationships that suggest critical thresholds in drought stress responses. Future studies may benefit from evaluating these thresholds among diverse accessions of switchgrass and other C4 grasses. mRNA profiles of leaf tissue from clonal replicates at various time points during drydown and recovery were generated by deep sequencing 3' mRNA tags using SOLiD.
Project description:In light of the changes in precipitation and soil water availability expected with climate change, understanding the mechanisms underlying plant responses to water deficit is essential. Toward that end we have conducted an integrative analysis of responses to drought stress in the perennial C4 grass and biofuel crop, Panicum virgatum (switchgrass). Responses to soil drying and re-watering were measured at transcriptional, physiological, and metabolomic levels. To assess the interaction of soil moisture with diel light:dark cycles, we profiled gene expression in drought and control treatments under pre-dawn and mid-day conditions. Soil drying resulted in reduced leaf water potential, gas exchange, and chlorophyll fluorescence along with differential expression of a large fraction of the transcriptome (37%). Many transcripts responded differently depending on time of day (e.g. up-regulation pre-dawn and down-regulation mid-day). Genes associated with C4 photosynthesis were down-regulated during drought, while C4 metabolic intermediates accumulated. Rapid changes in gene expression were observed during recovery from drought, along with increased water use efficiency and chlorophyll fluorescence. Our findings demonstrate that drought responsive gene expression depends strongly on time of day and that gene expression is extensively modified during the first few hours of drought recovery. Analysis of covariation in gene expression, metabolite abundance, and physiology among plants revealed non-linear relationships that suggest critical thresholds in drought stress responses. Future studies may benefit from evaluating these thresholds among diverse accessions of switchgrass and other C4 grasses.